A method in accordance with a preferred embodiment is based on the use of a microdroplet for containing the sample to be observed, or itself constituting said sample. The article by A B Theberge et al, “Microdroplets in Microfluidics: An Evolving Platform for Discoveries in Chemistry and Biology”, Angew Chem Int Ed 2010, 49, 5846-5868, presents a review of microfluidics techniques based on the use of liquid microdroplets for a variety of applications in chemistry and in biology. The term “microdroplets” means droplets with a diameter (or, more generally, principal dimension) in the range 1 μm to 100 μm.
Document US 2005/0221339 describes a method for the optical detection of chemical components in microdroplets manipulated in a microfluidics system.
The article by B Rotman, “Measurement of activity of single molecules of β-D-Galactosidase”, Proceedings of the Natl Acad Sci USA, 1961, 47, pp 1981-1991, describes a method for the detection of isolated molecules of an enzyme. In that method, an aqueous solution containing the enzyme (highly diluted) and a fluorogenic substrate for that enzyme is sprayed into a small volume of silicone oil confined between two microscope slides. Given that the oil is less dense than water and immiscible therewith, the solution forms microdroplets that are deposited on the upper surface of the lower microscope slide. Next, the droplets are analyzed by microfluorimetry with the aid of a microscope. The droplets containing at least one molecule of the enzyme appear as luminous disks, while the droplets containing none appear in the form of luminous rings.
The principal limitation to such methods resides in the low intensity of the light emitted by fluorescence. Similar problems arise when the light that is diffused (elastic or Raman diffusion) by an object of microscopic dimensions contained in a droplet is to be observed.